Project Summary Liver fibrosis is the consequence of chronic liver diseases, such as hepatitis B and C infection, and alcoholic and non-alcoholic steatohepatitis (NASH). Currently there is no effective anti-fibrotic agent for liver cirrhosis. Therefore, the unmet medical needs for liver fibrosis/cirrhosis are significant. Liver inflammation is a crucial mechanism for activation of hepatic stellate cell (HSCs) and liver fibrosis. Activated HSCs produce extracellular matrix (ECM) including collagen, fibronectin, and hyaluronic acids (HA). Endogenous HA have been implicated in the disease progression of lungs, kidneys, joints, heart, and brain. In the liver, patients with liver cirrhosis show elevated HA levels in the blood. However, the biological functions of endogenous HA produced in liver fibrosis have not been determined. HA are produced in high molecular weight forms (HMW; MW>1000kDa) by hyaluronan synthase (HAS) 1-3. In the setting of inflammation, HA are degraded into low molecular weight (LMW) forms (MW~100-300kDa). LMW-HA exacerbates tissue injury and inflammation through binding to its receptors, CD44 and TLR4. The objective of this study is to determine the biological functions of endogenous HA and its downstream effector pathway in HSC activation and in liver fibrosis. Based on previous publications and our preliminary studies, we hypothesize that HAS2-mediated HSC-derived HA and HA's downstream effector pathways promote HSC activation and liver fibrosis. Moreover, we further hypothesize that by targeting HA a new interventional strategy for treating liver fibrosis can be developed. To test our hypotheses, we will investigate if HAS2-mediated HA production in HSCs promotes liver fibrosis through loss- and gain-of-function approaches using HSC-specific Has2 knockout (Has2?HSC) mice and HAS2 transgenic (ASMA-HAS2 Tg) mice. We will also use human tissue samples to examine HSCs as the source of HAS2 and HA in human cirrhotic livers (Aim 1). We will investigate the transcriptional and posttranscriptional regulation of HAS2 expression in HSCs as the molecular mechanisms of dysregulated HAS2 expression in liver fibrosis (Aim 2). We will then investigate the role of Notch1 signaling as the HA's downstream effector pathway for HSC activation and liver fibrosis (Aim 3). Finally, we will examine interventional potential of blocking HA synthesis for treating liver fibrosis (Aim 4).